Mike Meade

Dr. Srinivasan

Developmental Biology

26 April 2015

The Development of Cardiogenic Mesoderm

Introduction

As already known, the three germ layers that develop during gastrulation

eventually develop into necessary structures for the body through the process of

organogenesis including vital organs such as the heart. What is to be explained in this

review is the specification and development of cardiac mesoderm, also known as the

heart field. The heart field is divided in the primary and secondary heart field. The

secondary heart field forms at the anterior portion of the body and the right ventricle and

cardiac outflow tract are eventually derivatives of it (Brand 2003). There are many

signaling paths and proteins that are involved in the formation of cardiac mesoderm

including BMP, noggin/chordin, wnt, mesp1, and mmp-3. Specifically there will be a

focus on BMP and noggin in this review as well as their interactions with all of the

aforementioned proteins and pathways.

The Importance of Inhibition

Before going any further it may useful to mention and briefly explain inhibition

since it is one of the most important aspects of development in regard to cardiac

mesoderm that first must be understood is the interaction of proteins and pathways in

their regard to turn others on or inhibit others. A very crucial example of this is the

ability of noggin and chordin to inhibit BMP, specifically BMP-4. By noggin and chording

inhibiting BMP the cardiac and blood-forming fields will not form in the center of an

embryo. Another inhibitory effect is the effect of dickkopf to repress wnt. An over

expression of wnt without inhibition leads to an overabundance of cells marked for blood

rather than cardiac mesoderm. Of course these paths of inhibition are not so simple

requiring cascading of signals and will be explained further later in the review.

Cardiogenic Mesoderm Develops from Lateral Plate Mesoderm

Cardiogenic mesoderm specifically develops from lateral plate mesoderm

through various signaling. While noggin and BMP are both important in developing

cardiogenic mesoderm, Wnt is also necessary and must be inhibited in order for later

signals of BMP to perform their job. BMP activates two genes known as Nkx2-35 and

Mesp1. Mesp1 is important for multiple reasons including that it prevents heart

progenitors from being re-specified as another type of mesoderm and also represses

brachyury, sox17, and goosecoid insuring that cardiac precursor cells will not become

other structures such notochord or endoderm (Gilbert p.453). In relation to Wnt, Mesp1

activates the dickkopf gene, which is a Wnt inhibitor. Wnt must be inhibited in order for

BMP to properly perform its function and signaling path.

PARM1 enhances BMP signaling

At this point it is obvious to see that BMP signaling is important in the

specification of cardiogenic mesoderm, but there also has to be signaling that regulates

BMP signaling. An article from 2012 showed the effects off the protein PARM1 on BMP

signaling. At normal levels PARM1 may not have as much of an effect on BMP, but

over expression enhances the effects of BMP2 and BMP4 (Nakanishi, N., Takahashi,

T., Ogata, T., Adachi, A., Imoto-Tsubakimoto, H., Ueyama, T., & Matsubara, H. 2012).

The expression of PARM1 is followed by mesodermal markers, which happens during

the differentiation of cardiomyocytes. Cardiomyocytes develop from cardiac mesoderm,

so the larger picture in BMP signaling and PARM1 is through the specification and

development of cardiac mesoderm to eventually develop fully into cardiomyocytes or

cardiac muscle cells.

Noggin Increases the Expression of MMP-3

As previously stated, noggin is an inhibitor of BMP and also critical during

development of cardiac mesoderm and cardiomyocytes. In addition to interacting with

BMP, noggin also interacts with mmp-3 by increasing the expression of it. Without

noggin present BMP is an inhibitor of mmp-3. If mmp-3 is inhibited by BMP there is a

decrease in cardiac differentiation. Mmp-3 is necessary for the activation of genes such

as Gata-4 which are specific cardiac markers that allow for cells to differentiation and be

specified for cardiac development (Hong, S., Kang, J., Park, J., Ryu, E., Choi, S., Lee,

S., . . . Seo, J. 2010). In the development of cardiac mesoderm and cardiomyocytes it

is often important to not only understand how certain proteins and genes are turned on

and what they contribute to the cascading of signals, but also to understand why some

are turned off. In this part of development it is necessary for BMP to be inhibited by

noggin. If it were the case that noggin was not inhibiting BMP, then mmp-3 would not

be able to activate Gata-4 and other cardiac markers. Without the activation of Gata-4,

differentiating cells would not be able to be marked for creating cardiac mesoderm and

then developing into cardiomyocytes.

BMP Activates MESP1

As noted earlier, wnt signaling must be inhibited by genes such as dickkopf and

Cerberus in order for BMP to convert lateral plate mesoderm to cardiac mesoderm. If

BMP is not present in order to activated mesp1, which activates dickkopf, then wnt

would not be inhibited. Wnt, specifically wnt8 actually inhibits the formation of cardiac

mesoderm, but instead is important in specifying cells to the fate of blood. While blood

is clearly a very important and necessary part of the body of nearly all living beings, if

wnt is never inhibited then there can be no heart formation because the formation of

cardiac mesoderm and cardiomyocytes may be interrupted and possibly not form all

due to over expression of cells marked for the fate of blood. This is another case in

which is critical to see how inhibition of certain signaling paths is required for other

structures to form. Over expression of wnt8 without ever being inhibited would convert

all lateral plate mesoderm to become specified for blood and cardiac mesoderm would

not be able to properly develop.

Current Model of Cardiogenic Mesoderm Development

The current model of the development of cardiogenic mesoderm is tied in with all

of the aforementioned signaling pathways as well as later plate mesoderm. Wnt, BMP,

noggin, Mesp1, and others are all crucial. The neural tube secretes wnt signals that

specify lateral plate mesoderm to hemangiogenic mesoderm (blood). In the anterior

part of the body, dickkopf inhibits wnt, which allows BMP and FGF8 to work on

converting lateral plate mesoderm to cardiogenic mesoderm. Also to be noted here is

that FGF and BMP are both needed in the posterior for heart formation (Brand 2003)

Chordin and noggin are secreted from the notochord so that blood-forming fields are not

formed in the middle of the embryo. Chordin and noggin of course are inhibitors of BMP

and block BMP signaling. BMP activates Mesp1, which in turn activates dickkopf.

Mesp1 when activated also prevents genes that would specify cells for fates other than

cardiac mesoderm. This is the basic model of the development and specification of

cells for cardiac mesoderm. To go further into the development of cardiomyocytes is to

further understand BMP signaling and its inhibition by noggin. BMP must be inhibited

by noggin in order for mmp-3 to activate cardiac markers such as gata-4, which mark

differentiating cells for the creation and development of cardiomyocytes.

The Problems With the Current Model

The current model of the specification of cardiogenic mesoderm and the

development of cardiomyocytes is still not perfect and an area of current research. One

thing that is unclear is how noggin can elevate mmp-3 expression (Hong, S., Kang, J.,

Park, J., Ryu, E., Choi, S., Lee, S., . . . Seo, J. 2010). In the experiment entitled

“Association of Matrix metalloproteinase-3 with cardiogenic activity during Noggin-

induced differentiation of mouse embryonic stem cells” from 2010 the researchers

explored how inducing noggin affects mmp-3. They were able to show that mmp-3 is

regulated and has increased activity due to noggin, but were unable to tell exactly how

noggin was doing this. Also it seems unclear if BMP is alone required to activate mesp1

and if another signaling path could achieve the same thing. This is important because it

leads to the inhibition of wnt8. This leads to another gap in the model. If nothing is

there to inhibit wnt, then can cardiac mesoderm even attempt to develop or do all cells

become specified for hemangiogenic mesoderm? It is crucial to understand all of these

aspects of development in order to understand if each signaling pathway is sufficient for

their role and if their over expression (in other words not being inhibited when

necessary) causes serious issue in development.

Possible Future Research and Experiments

Certain experiments would be necessary in order to fill the gaps in the current

models for the development of cardiac mesoderm and cardiomyocytes. An experiment

to attempt to solve the gap of how noggin elevates mmp-3 could be to see how BMP

acts on mmp-3 in the absence of noggin. In other word perform a “lose it” experiment

losing noggin. Also it would be necessary to ensure that BMP was close enough to act

on mmp-3. Next is the question of whether the absence of BMP would cause over

expression of wnt where it is normally inhibited and if cardiac mesoderm in those areas

(posterior) would be able to form. This could be another “lose it” experiment in which

BMP was removed from the posterior. If cardiac mesoderm was still able to form in the

posterior half, then wnt8 may not be capable of inhibiting it on its own or another

signaling pathway may be able to activate mesp1, which would in turn activate dickkopf

to block wnt signaling. If cardiac mesoderm was not able to develop and there was an

overabundance of cells being specified for hemangiogenic mesoderm then the opposite

would be true.

Annotated Bibliography

1. Nakanishi, N., Takahashi, T., Ogata, T., Adachi, A., Imoto-Tsubakimoto, H.,

Ueyama, T., & Matsubara, H. (2012). PARM-1 promotes cardiomyogenic

differentiation through regulating the BMP/Smad signaling pathway. Biochemical

and Biophysical Research Communications,428(4), 500-505.

2. Hong, S., Kang, J., Park, J., Ryu, E., Choi, S., Lee, S., . . . Seo, J. (2010).

Association of Matrix metalloproteinase-3 with cardiogenic activity during Noggin-

induced differentiation of mouse embryonic stem cells. International Journal of

Cardiology, 141(1), 49-60.

3. Kattman, S., Witty, A., Gagliardi, M., Dubois, N., Niapour, M., Hotta, A., . . .

Keller, G. (2011). Stage-Specific Optimization of Activin/Nodal and BMP

Signaling Promotes Cardiac Differentiation of Mouse and Human

Pluripotent Stem Cell Lines. Cell Stem Cell, 8(2), 228-240.

4. Liu, Y., Asakura, M., Inoue, H., Nakamura, T., Sano, M., Niu, Z., . . . Schneider,

M. (2007). Sox17 is essential for the specification of cardiac mesoderm in

embryonic stem cells. PNAS,104(10), 3859–3864-3859–3864.

5. Schneider, V., & Mercola, M. (2001). Wnt antagonism initiates cardiogenesis in

Xenopus laevis. Genes and Development, 15(1), 304-315.

6. Brand, T. (2003). Heart development: Molecular insights

into cardiac specification and earlymorphogenesis. Developmental

Biology, 258(1), 1-19.

7. Gilbert, S. (2014). Lateral Plate Mesoderm and the Endoderm.

In Developmental Biology(10th ed., Vol. 1, pp. 451-453). Sunderland, MA:

Sinauer Associates.

Figures/Examples

Figure 1: Example of BMP being antagonized (inhibited) in blastula stage. While this is

not during gastrulation as discussed in the review it allows for the ability to see how

BMP is inhibited and its expression patterns.

Figure 2: Development pathway to cardiac mesoderm showing BMP gradients

Figure 3: Lateral Plate Mesoderm to Cardiac Mesoderm Specification